熊蜂ISSR-PCR体系的建立与优化及其在亲缘关系分析中的应用

以小峰熊蜂Bombus hypocrita Pérez为实验材料, 建立并优化简单序列重复区间 (inter-simple sequence repeat, ISSR)扩增多态反应体系, 并运用ISSR标记和NTSYS聚类软件分析群内亲缘关系和亚家系组成。结果表明: 在10 μL的PCR反应体系中各组分的适宜终浓度为1×PCR 缓冲液、0.3 mmol/L 反应底物、1.25 μmol/L 引物、3 mmol/L 镁离子、2.5 U Taq 酶和8.18~23.76 μg/mL DNA模板。用此优化后的反应体系对小峰熊蜂基因组DNA进行ISSR-PCR扩增, 能有效分析群内亲缘关系和亚家系组成。

Establishment and optimization of ISSR-PCR reaction system and its application in the genetic coefficient analysis of Bombus hypocrita Perez(Hymenoptera: Apidae)

In this study, the ISSR-PCR reaction system was established, optimized and then used to analyze the genetic coefficient and the subfamily composition of Bombus hypocrita Pérez.The optimized reaction system was established as follows: 1×PCR buffer, 0.3 mmol/L dNTP, 1.25 μmol/L primer, 3 mmol/L Mg2＋, 2.5U Taq polymerase and 8.18-23.76 μg/mL DNA. Our results indicated that ISSR markers obtained with this optimized system could be effectively used to analyze the genetic coefficient and subfamily composition.